P-(2-(1-phenyl-3,5-dioxy-4-n-butyl)-pyrazolydinyl)phenylphosphoric acid,its organic and inorganic salts

ABSTRACT

P-(2-(PHENYL -3,5- DIOXY - 4 - N-BUTYL)-PYRAZOLYDINYL) PHENYLPHOSPHORIC ACID OF THE FORMULA   1-(4-((HO-)2-P(=O)-O-)PHENYL),2-PHENYL,4-(CH3-(CH2)3-)-   IMIDAZOLIDINE-3,5-DIONE   AND PHARMACEUTICALLY ACCEPTABLE INORGANIC AND ORGANIC SALTS THEREOF ARE INCLUDED IN THIS INVENTION. THE COMPOUNDS HAVE ANTIPHLOGESTIC AND ANTIPYRETIC ACTIVITY.

United States Patent 4 Claims ABSTRACT OF THE DISCLOSURE p-[Z-(phenyl 3,5 dioxy 4 n-butyl)-pyrazolydinyl] phenylphosphoric acid of the formula and pharmaceutically acceptable inorganic and organic salts thereof are included in this invention. The compounds have antiphlogestic and antipyretic activity.

SUMMARY OF INVENTION This invention refers to the p-[2-(1-phenyl-3,5-dioxy- 4-n-butyl)-pyrazolydinyl]-phenylphosphoric acid, its organic and inorganic salts and the process to prepare them DETAILS OF INVENTION In particular this invention refers to p-[2-(phenyl-3,5- dioxy-4-n butyl)-pyrazolydinyl]phenylphosphoric acid of Formula I H;CHzCH2-CH:

and to the process to prepare the same. This invention relates also to the salts formed by this acid and pharmaceutically acceptable inorganic or organic bases. Among the possible inorganic salts there are sodium salt, po-

tassium salt and calcium salt, whereas among the organic ones are included those salts formed by basic substances having pharmacological activity such as u-p-phenetidino- N-n-propylpropionamide, aminopyrine, ethanolamine and diethanolamine.

For a systematic study of the pharmacological activities, the calcium salt of Formula II CHg-CHg-CHg-CH; 2 (II) in which x is from 0 to 14, and the sodium salt of Formula -III "ice in which x is from 0 to 8 were chosen. They are stable in aqueous solution or in bulfers with dilferent pH. More particularly, the sodium salt, at room temperature in buffer with pH 8.8, does not show degradation after 20 hours; whereas at C. in buffer solution (pH 8.8), there is 50% of degradation. The sodium salt III is easily dissolved in Water and in buffers with pH 6.5-7 up to a concentration of about 500 mg./ml. For this reason it can be administered intramuscularly or intravenously at high concentration. When the calcium II and sodium III salts were administered orally, intramuscularly, intravenously or rectally, no intolerance of the drug was noticed.

Furthermore, the salts of this invention are not absorbed in the stomach, but they pass directly into the intestine where they are absorbed. Therefore they have no irritant effect on the gastric mucous membrane such as those caused by other drugs.

The compounds of this invention are prepared by reacting 4-n-butyl-2-(p-hydroxyphenyl)-1-phenyl-3,5-pyrazolydindione with phosphoryl chloride dissolved in an aprotic solvent such as ethyl ether, benzol, toluol, xylol, tetrahydrofuran, petroleum ether or other solvent stable under reaction conditions. To neutralize the hydrochloric acid split off during the reaction, the solvent is added to an organic or inorganic base able to form salts with hydrochloric acid. In the process of the invention anhydrous pyridine was used, but also trialkylamine, dialkylary1- amines, N'-alkylazocycloalkane, inorganic oxides, hydroxides, carbonates or bicarbonates are also suitable.

The reaction temperature ranges between 0 C. and 50 C. The reaction product is treated according to the well known methods of the preparatory organic chemistry. The cooled mixture is added with water, then it is dried and the residue is neutralized by aqueous MgO. The solution is desalted using an ion exchanger resin, such as Amberlite I.R.120, or purified by chromatography over a resin such as Dowex-L00 To isolate the calcium salt it proved more advantageous to avoid using resins. The reaction mixture is filtered when cold and the precipitate is dissolved, at room temperature, in water having a highly alkaline pH which was obtained by adding CaO and/or Ca(OH) The solution thus obtained is concentrated to a small volume and added to a Water miscible solvent such as methyl alcohol, ethyl alcohol, acetone or other solvent in which the salt is insoluble. The precipitate obtained is the calcium salt of Formula II which can be further purified by dissolution in water and precipitation by adding methyl or ethyl alcohol or acetone. Generally, to obtain an analytically pure salt it is suflicient to repeat this operation twice. The calcium salt absorbed crystallization water. After five days of exposure to the air a salt with 14 molecules of crystallization water was isolated.

Additionally, salts with a different number of water molecules were isolated and among them a salt with 8 molecules of crystallization water was chosen for the pharmacological tests. The salt was obtained leaving the reaction product in the air contact for 7 hours, drying it at 70 C. and under high vacuum.

The anhydrous calcium salt or hydrate form of it is dissolved in 50-100 volumes of distilled 'water and the solution is poured on an ion exchanger resin column such as Amberlite I.R. or Dowex 50 W X2. The eluted product is collected when the pH is clearly acid to pH between 1 and 5. From this solution we obtain:

(1) the free acid by concentrating the solution under vacuum at a temperature lower than 45 C. until 21 volume equal to one tenth of the starting solution and then by lyophilizing it;

(2) the sodium salt by adding a solution of N NaOH, or of N Na CO or of N NaHCO to reach pH 7.8-8, and

3 concentrating under vacuum at a temperature lower than 45 C. and lyophilizing; (3) the addition salt with an organic base dissolved in a water miscible solvent such as methanol, ethanol,

TABLE 4 Carrageenan oedema on adrenaleetomized rats Percent inhibition in diflerent times acetone: the solution is concentrated under vacuum r Number of at a temperature lower than 45 C., the residue is Mg-lkg-dose 311011 95 6110MB animals washed several times with a water non-miscible solvent 24 2g such as ethyl ether, benzol, and dissolved in a little 3 1-3: fi-gg i8 acetone; by settling at a temperature between 0' C. and 10 C. a precipitate is obtained which is purified 10 by several washings with a water non-miscible solvent. TABLE 5 The chemical structure of the compounds are confirmed Fmmu" mites nmnal by infrared and ultraviolet spectra, by the correspondence Perceragtinhibition between calculated values for the elements and values gr Number of found by chemical analysis, and by the results of the s-l ge 4hours Shours animals enzymatic hydrolysis which produces the starting com- 5+5 pounds. 23.40 -1s.33 5+5 The octohydrate calcium salt (II) and the sodium salt 5+5 (III) of (I) acid were pharma-cologieally tested to evaluate the acute toxicity, the antiphlogistic and antipyretic TABLE 6 activity. During these tests no phenomena of intolerance Formaun mites on adrenalectomized rats of the two drugs was exhibited. In the test of acute toxic- 1 h 1ty the substance 'was administered in differing doses, gi gl spaced on logarithmic scale, in groups of 5 animals each M d 4h h Number n dose. Mortality was observed for 8 days and the LD use Ours 8 outs mums was calculated according to the Spearman-Karber method 3%; 33- 3 (ref. Finney, D. 1.: Statistical Method in Biological Asj 46:24 5+5 say, 2nd ed., London (1964), page 524) TOXICITY TABLE 1 LD mgJkg. (LF, P=0.05)

Mice Rats Compound Os I.v. Os Lv. Lm.

(Ia-H1O salt 1,117(9431,226) Sodium salt 401(311-517) 170055-185) 782(669-924) 205(1343l6) 213(172-263) The antiphlogistic activity was investigated using the 01- TABLE 7 lowmg tests: Granuloma pouch on normal rats (1) Qarrageenan Oedema-Winter C. A. et coll.: Proc. Percent inhibition Soc. Exper. Biol. Med. 111, 544 (1962) r Pouch Number of (2) Formalin Ascite-Wilhelmi G.--Schwz. Med. Wshr. MgJkg. dose Exudate weight animals 185 (1953) -32.59 aaaz 5 (3) Granuloma pouch-Selye H. Proc. Soc. Exp. B101. 3.9% :s zggg 2 Med. 82, 328 (1953).

In Table 2 there are data referring to the octahydrate 50 TABLE 8 calcium Salt- Granuloma pouch on adrenaleetomized rats Percent inhibition BLE 2 Pouch Number of oarmgeenan oedema (rat) 5 5 MgJkg. dose Exudate weight animals Percent reduction Mg./kg.los dose of the oedema g -27.27 -6.6 5 -11.9 -1s.41 -21.s3

In Tables 3 to 8 there are results referring to the sodium salt.

TABLE 3 Carrageenan oedema on normal rats Percent inhibition in difierent times Number 01 MgJkg. dose ahours 43.6 hours Ghours animals The antipyretic activity was tested in the octahydrate calcium phosphate (1) salt and the results are in Table 9.

TABLE 9 antipyresis-rat, 0s

Percent reduction of temperature MgJkg. dose TABLE 10 After After After Number of MgJkg. dose 1 hour 2 hours 4 hours animals The phosphate, the object of this invention in the form of free acid, or as sodium or calcium salt, or as addition salts with organic compounds, can be administered separately or associated with other substances having therapeutic activity, orally, intramuscularly, intravenously, or rectally at a dose between 50 and 2000 mg./kg. daily.

For the oral administration, tablets can be prepared according to the following composition:

For suppositories:

Bisodium (III) salt fat excipient enough for 1.5 g. 50 Bisodium (III) salt fat excipient enough for While for intramuscular or intravenous syringes:

Bisodium (III) salt lyophilized 5O Distilled water or physiologic solution, 1 ml.

The following examples are illustrative of the invention and do not limit the invention.

EXAMPLE 1 p-[2-(1-phenyl-3 ,5 -dioxy-4-n-butyl -pyrazolydinyl] phenylphosphate calcium salt g. (0.066 mole) of phosphoryl chloride just distilled and 350 ml. of ethyl ether are put in a three necked flask. 20 g. (0.062 mole) of 4-butyl 2 (p-hydroxypheuyl)-1- phenyl 3,5 pyrazolydindione and 20 ml. of anhydrous pyridine dissolved in 250 ml. of anhydrous ether are slowly added to the solution under stirring and at room temperature. The precipitate formed is filtered, washed with ethyl ether, then dried under vacuum: yield 40 g.

The solid, washed with water, is suspended in 1,800 ml. of saturated calcium hydroxide solution; after the substance has been dissolved, calcium oxide is added until the solution has pH 12, then it is filtered. The ethereal solution obtained by washing the evaporated precipitate gives an oily residue which is treated with a saturated solution of calcium hydroxide and then with calcium oxide to reach pH 12, then it is filtered. The two aqueous solutions collected are concentrated under vacuum and at 35 C.

to a final volume of 300 ml. By adding 300 ml. of ethanol this solution gives a precipitate which is washed with ethyl alcohol and ethyl ether. Yield: 18.5 g.

The precipitate is suspended in 500 ml. of water and kept under stirring for 2 hours. It is filtered, the insoluble residue is discarded whereas the solution (pH 12.1) is concentrated under vacuum and at 35 to a volume of 250 ml. 300 ml. of ethyl alcohol are added to the solution. The insoluble residue is filtered, washed with a hydroalcoholic solution (1:2) then with ethanol and finally with ethyl ether. The product obtained (10.8 g.) is further purified by extracting with 300 ml. of water. The insoluble residue is discarded and the aqueous solution (pH 9.3) is concentrated to ml.; by adding 150 ml. of ethanol, a precipitate is obtained which is washed with an hydroalcoholic solution 1:2), then ethyl alcohol, then ethyl ether, and then it is dried under vacuum. Yield: 6.6 g. of p-[2-(1-pheny1 3,5 dioxy 4 n butyl)- pyrazolydinyl]phenylphosphate calcium salt.

The calcium salt left in contact with the air absorbs crystallization water; after 7 hours a salt with 8 H O is obtained which is dried under high vacuum and at 7 0 C After 5 days the water molecules are 14.

(a) Elementary analysis (with dried sample under high vacuum and at 70 C.).Calculated for aa 3s 4 12 a 2' 2 (percent): C, 42.77; H, 4.87; N, 5.25; P, 5.81; Ca, 11.27. Found (percent): C, 42.57; H, 5.15; N, 5.24; P, 5.80; Ca, 11.41.

H O 13.5 (K.F.) U.V

M m e=3 1,300 (pH 1; in HCl 0.1 N) A =262 m e=39,800 (pH 8.8; borated buffer +HC1 0.1 N) LR. (KBr): 1620; 1310; 1240; 1010 ems- Electrophoresis: Munketell paper n. 20acetate buifer pH 5.2; one spot only migrates toward the visible anode at UN.

Calculated for C H N O Ca P -14H O (percent): C, 38.81; H, 5.49; N, 4.76; Ca, 10.20; P, 5.60. Foind (percent): C, 38.67; H, 5.86; N, 4.43; Ca, 10.52; P, .60.

A =235 m t; -e=30,800 (pH 1; HCl 0.1 N) M :262 m e=40,500 (pH 8.8)

IR. (KBr): 1620; 1310; 1240; 1010 cm.-

Enzymatic hydrolysis p-[2-(1-phenyl-3,5-dioxy-4-n-butyl)- pyrazolydmyl]phenylphosphate calcium salt 29.43 mg. of 4-butyl-2-(p-hydroxyphenyl)-1-phenyl-3, S-pyrazo'lydindione phosphate calcium salt 8 H O dissolved in 100 ml. of citrate buffer pH 4.8 are added to a solution of 80 mg. of acidulous phosphatase (Mann Research Laboratories) dissolved in 20 ml. of distilled H O. The mixture is left at room temperature for 45 minutes; the hydrolysis is controlled while determining the inorganic phosphorus which splits off. The solution is extracted with chloroform which is washed with water to neutralization. It is dried over Na SO and the solvent is discarded under vacuum. The residue is p-hydroxyphenylwater, the mixture is filtered. The solution is poured on a Dowex 50 W X2 H+ (50-1-100 mesh) column (5.5 cm. diameter, 30 cm. height) and it is eluted with distilled water. The eluted solution having a pH lower than 4 is collected. 1,200 ml. of solution were collected, then concentrated under vacuum at a temperature lower than 40 C. to a volume of 60 ml. This filtered solution is lyophilized. Yield: 3.2 g. (80%) of p-[2-(1-pheuyl-3,5-dioxy-4- n-butyl)pyrazolydinyl]phenylphosphate acid.

Calculated for C H N O P (percent): C, 56.44; H, 5.23; N, 6.92; P, 6.67. Found (percent): C, 55.55; H, 5.59; N, 7.00; P, 7.15.

U.V in HCl 01 N:

k =235 mu, e=15,l00; in butler pH 8.8:

)l =260,u., e=19,450.

EXAMPLE 3 p- [2-( 1-pheny1-3,5 -dioxy-4-n-butyl) pyrazolydinyl] phenylphosphate bisodium salt 12 g. of calcium p-[2-(l-phenyl-3,5-dioxy-4-n-butyl)- pyrazolydinyl]phenylphosphate are dissolved in 600 ml. of water and filtered. The solution is poured on a Dowex 50 W X2 H+ (50+100 mesh) column (3.5 cm. diameter, 30 cm. height) and it is eluted with distilled water. The eluted solution having pH lower than 4 is collected. 1200 ml. of solution are collected and, by adding N NaOH, its pH reaches 6.8-7 with N NaOH. The solution is concentrated under vacuum and at a temperature lower than 40" C. to a volume of 60 ml. This solution is filtered and 13 0- philized. Yield: 3.3 g. of p-[2-(l-phenyl-3,5-dioxy-4-nbutyl)pyrazolydinyl]phenylphosphate bisodium salt.

Calculated for C H N O PNa -4H O (percent): C, 43.48; H, 5.22; N, 5.38; P, 5.95. Found (percent): C, 43.43; H, 4.92; N, 5.55; P, 5.65. E 0: 14.8% (calculated 13.9%

EXAMPLE 4 p-[2-(1-phenyl 3,5 dioxy 4 n-butyl)pyrazolydinyl] phenyllphosp'hate of a p phenetidino-N-n-propyl-propionamide 9.5 g. of p-[2-(1-phenyl-3,5-dioxy-4-n-butyl)-pyrazolydinyl]phenylphosphate calcium salt are chromatographed on Dowex 50 X2 form 11* by eluting with water. 1000 ml. of solution having pH lower than 4 are collected. The eluted mixture is mixed with an ethanolic solution of a-pphenetidino-N-n-propyl-propionamide (8.04 g.) and the solution obtained is evaporated to dryness at lowered pressure and at 40 C. The gummy residue obtained is washed three times with 60 cc. of ethyl ether each time. The residue is then diluted in 70 ml. of acetone and the solution is left in the refrigerator. The crystallized prodnet is washed with ethyl ether. After drying under high U.V.z M :240 ma; e=30,600.

We claim:

1. p-[2 (l-phenyl-3,5-dioxy-4-n-buty1)-pyrazolydinyl] phenylphosphoric acid having the formula:

HrCHr-CHr-Cih and its pharmaceutically acceptable organic and inorganic salts.

2. A calcium salt of the acid of claim 1 having the formula:

in which x may be from 0 to 14.

3. A sodium salt of the acid of claim 1 having the formula:

Na 0 ONa Care H20 CHz-CHg-CHz-CH;

in which .x may be from 0 to 8.

4. p- [2 1-phenyl-3,5-dioxy-4-n-butyl pyrazolydinyl] phenylphosphate of a p-phenetidino-N-n-propyl-propionamide.

References Cited UNITED STATES PATENTS 3,629,282 12/1971 Negrevergne 260-310 B 3,703,528 11/ 1972 Casadio et a1 260-310 B HENRY R. JILES, Primary Examiner G. T. TODD, Assistant Examiner US. Cl. X.R. 424-273 

